LOCUS       sequence-249189-        5007 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Enhancer reporter construct containing the chicken EdnrB enhancer, 
            E2 driving Citrine activity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5007)
  AUTHORS   Ling ITC, Sauka-Spengler T
  TITLE     Early chromatin shaping predetermines multipotent vagal neural crest
            into neural, neuronal and mesenchymal lineages.
  JOURNAL   Nat Cell Biol. 2019 Dec;21(12):1504-1517. doi: 
            10.1038/s41556-019-0428-9. Epub 2019 Dec 2.
  PUBMED    31792380
REFERENCE   2  (bases 1 to 5007)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41556-019-0428-9"; journalName: "Nat Cell Biol"; date: 
            "2019-12"; volume: "21"; issue: "12"; pages: "1504-1517"
COMMENT     Sequence Label: pTK-EdnrbE2-Citrine
FEATURES             Location/Qualifiers
     source          1..5007
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        53..198
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          276..408
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     391..410
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      462..471
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(468..470,471..473,474..1184)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(513..534)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(774..793)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1121..1142
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1356..1477
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1367..1386)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1421..1440
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1725..1742)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1896..2484)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1976..1995)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2655..3446,3447..3515))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3278..3297
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3516..3620)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3647..4102
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3734..3753)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3944..3965
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4233..4281
                     /note="synthetic polyadenylation signal"
     misc_feature    4295..4386
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4335..4354
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 tttgctggcg gtgtccccgg aagaaatata tttgcatgtc tttagttcta tgatgacaca
       61 aaccccgccc agcgtcttgt cattggcgaa ttcgaacacg cagatgcagt cggggcggcg
      121 cggtcccagg tccacttcgc atattaaggt gacgcgtgtg gcctcgaaca ccgagcgacc
      181 ctgcagcgac ccgcttaaca gcgtcaacag cgtgccgcag atcccaagct tgcggccgct
      241 taagctgcag aagttggtcg tgaggcactg ggcaggtaag tatcaaggtt acaagacagg
      301 tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg cgtttctgat
      361 aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt gtccactccc
      421 aggttcaata cagctcttaa gcggccgcaa gctagcttgc cgccaccatg gtgagcaagg
      481 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
      541 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
      601 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccacct
      661 tcggctacgg cctgatgtgc ttcgcccgct accccgacca catgaagcag cacgacttct
      721 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
      781 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
      841 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
      901 actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga
      961 acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
     1021 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagctacc
     1081 agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
     1141 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtgagcc tctagactcc
     1201 ctcagcggcg cgcctggtgt tcctcgagat cggagtccta aatcgactgc cttagcggta
     1261 ggacagtaga tcagtcagcg acgtgacgaa ccctcgcagg atcaagatcc cacgtgtcta
     1321 gagtcggggc ggccggccgc ttcgagcaga catgataaga tacattgatg agtttggaca
     1381 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     1441 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     1501 tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa acctctacaa
     1561 atgtggtaaa atcgataagg atccgtcgac cgatgccctt gagagccttc aacccagtca
     1621 gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc acttatgact gtcttcttta
     1681 tcatgcaact cgtaggacag gtgccggcag cgctcttccg cttcctcgct cactgactcg
     1741 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     1801 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     1861 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     1921 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     1981 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     2041 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     2101 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     2161 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     2221 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     2281 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     2341 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     2401 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     2461 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     2521 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     2581 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     2641 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     2701 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     2761 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     2821 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     2881 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     2941 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     3001 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     3061 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     3121 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     3181 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     3241 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     3301 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     3361 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     3421 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     3481 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     3541 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     3601 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgc gccctgtagc
     3661 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     3721 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     3781 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     3841 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     3901 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     3961 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     4021 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     4081 aaaatattaa cgcttacaat ttgccattcg ccattcaggc tgcgcaactg ttgggaaggg
     4141 cgatcggtgc gggcctcttc gctattacgc cagcccaagc taccatgata agtaagtaat
     4201 attaaggtac gggaggtact tggagcggcc gcaataaaat atctttattt tcattacatc
     4261 tgtgtgttgg ttttttgtgt gaatcgatag tactaacata cgctctccat caaaacaaaa
     4321 cgaaacaaaa caaactagca aaataggctg tccccagtgc aagtgcaggt gccagaacat
     4381 ttctctatcg ataggtaccg agctcgagtt gccagtgctt gtaggaccgc tcaagtgctt
     4441 gcagctcaca attaccctgc tggcagttgt aaccgccgca ctaggactaa gtcgcttgtg
     4501 tcttgtcatt acaatgcttc cttttgtccg atggctgcaa atgcttgaca gtcattagaa
     4561 agtcatccaa ggaaatactt cctttccatg tacaagatgc ttgtagtgga tttttaacac
     4621 tgagctttct cagtaactca gatagcatta gcaaaagaat agtttcttac agcctgcctc
     4681 tgcccttaag gttcttttat aaagagaaca gaagctcttt gttgttactc cttcacacgt
     4741 tattaacaga ggtttaacag aaagaagagg ggcttgttat agtccgctgc aatttgtctt
     4801 tttcagttca ttcggatctc catcatgtcc aaccagaaga cggattaacc cagatccagg
     4861 gtggctgctt ccatactaaa cagaaaatga attttaaaaa gtcagacagt gtaatctctc
     4921 tctgccaaga acctgttggc tagcccgggc tcgagatctg cgatctaagt aagcttgctg
     4981 cttcatcccc gtggcccgtt gctcgcg
//
